September 2010
Volume 51, Issue 9
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Retina  |   September 2010
Evaluation of Macular Structure and Function by OCT and Electrophysiology in Patients with Vitelliform Macular Dystrophy Due to Mutations in BEST1
Author Affiliations & Notes
  • Patrik Schatz
    From the Department of Ophthalmology, Glostrup Hospital, University of Copenhagen, Denmark;
    the Department of Ophthalmology, Lund University Hospital, University of Lund, Sweden;
    the Gordon Norrie Centre for Genetic Eye Diseases, National Eye Clinic for the Visually Impaired, Kennedy Center, Glostrup, Denmark; and
  • Hanna Bitner
    the Department of Ophthalmology, Hadassah-Hebrew University Medical Center, Jerusalem, Israel.
  • Birgit Sander
    From the Department of Ophthalmology, Glostrup Hospital, University of Copenhagen, Denmark;
  • Stig Holfort
    From the Department of Ophthalmology, Glostrup Hospital, University of Copenhagen, Denmark;
  • Sten Andreasson
    the Department of Ophthalmology, Lund University Hospital, University of Lund, Sweden;
  • Michael Larsen
    From the Department of Ophthalmology, Glostrup Hospital, University of Copenhagen, Denmark;
  • Dror Sharon
    the Department of Ophthalmology, Hadassah-Hebrew University Medical Center, Jerusalem, Israel.
  • Corresponding author: Patrik Schatz, Department of Ophthalmology, Glostrup Hospital, 2600 Glostrup, Denmark; patrik.schatz@med.lu.se
Investigative Ophthalmology & Visual Science September 2010, Vol.51, 4754-4765. doi:10.1167/iovs.10-5152
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      Patrik Schatz, Hanna Bitner, Birgit Sander, Stig Holfort, Sten Andreasson, Michael Larsen, Dror Sharon; Evaluation of Macular Structure and Function by OCT and Electrophysiology in Patients with Vitelliform Macular Dystrophy Due to Mutations in BEST1. Invest. Ophthalmol. Vis. Sci. 2010;51(9):4754-4765. doi: 10.1167/iovs.10-5152.

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      © ARVO (1962-2015); The Authors (2016-present)

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Abstract

Purpose.: To analyze retinal structure and function in vitelliform macular dystrophy (VMD) due to mutations in BEST1.

Methods.: Patients from five Swedish and four Danish families were examined with electrooculography (EOG), full-field electroretinography (ffERG), multifocal ERG (mfERG), optical coherence tomography (OCT), and fundus autofluorescence photography (FAF). Genetic analysis of the BEST1 gene was performed by direct sequencing.

Results.: Mutations in BEST1 have been reported previously in the Swedish families. In the Danish families, four disease-causing missense mutations were found, one of which is novel: c.936C>A (p.Asp312Glu). The mutation was homozygous in a 9-year-old boy and heterozygous in his father in a consanguineous family. ffERG rod response was reduced in the homozygous boy, but normal in the heterozygous father. EOG was reduced in all but two patients and did not correlate with the ffERG results. OCT ranged from normal to cystoid edema and thickening of the outer retina–choroid complex. Decreased mfERG amplitudes, increased mfERG latencies, and loss of integrity of the foveal photoreceptor inner/outer segment junction, correlated with decreased vision. FAF demonstrated hyperautofluorescence beyond the ophthalmoscopic changes in several patients.

Conclusions.: The finding of a homozygous dominant mutation in a patient with VMD and evidence of widespread retinal degeneration may imply that the pathogenesis of the generalized retinal degeneration differs from that of the macular degeneration. A relative agreement between hyperautofluorescence by FAF, reduced retinal function, and VMD implies that the hyperautofluorescence emanates from lipofuscin and A2E. A potential therapy for VMD, involving the inhibition of the retinoid cycle, is suggested.

Human bestrophin-1 (BEST1, MIM 607854; Mendelian Inheritance in Man; National Center for Biotechnology Information, Bethesda, MD)–associated retinopathy has been the focus of several papers recently. The spectrum of retinopathies caused by mutations in this gene includes at least autosomal dominant (AD) Best vitelliform macular dystrophy (BMD), autosomal recessive (AR) bestrophinopathy (ARB), AD vitreoretinochoroidopathy (ADVIRC), AD and AR retinitis pigmentosa. 16 Moreover, we have recently reported a family with a rare BEST1 genotype in which two sisters aged 30 and 33 were compound heterozygous for a missense mutation and a null mutation, featuring vitelliform dystrophy and electrophysiological signs of widespread retinal degeneration, which may be similar to ARB. 7 In dogs, a specific retinopathy, termed canine multifocal retinopathy, has been associated with recessive mutations in best1. 8  
The various retinopathies associated with mutations in BEST1 may share some pathogenetic features. The protein product of BEST1, bestrophin, localizes to the basolateral plasma membrane of the retinal pigment epithelium (RPE). 9 It has been suggested to act as a Ca2+-sensitive chloride channel or as a modulator of voltage-sensitive Ca2+ channels in the RPE. 10,11 It has recently been shown that bestrophin channel activity is regulated by ceramide, a lipid-signaling molecule, through phosphorylation by protein kinase C (PKC) and dephosphorylation by protein phosphatase 2A at position serine 358. 12 It was suggested that ceramide accumulation in early retinopathy inhibits bestrophin function, leading to abnormal fluid transport across the retina. This mechanism may be a general pathogenetic one with a role in various retinopathies, not only those associated with mutations in BEST1
Furthermore, there is an increasing understanding of the genotype–phenotype correlations that may provide further insight into the pathogenesis of the various retinopathies associated with mutations in BEST1. For example, ADVIRC has been associated with mutations that affect mRNA splicing, whereas most cases of BMD are due to heterozygous missense mutations. 5 In AD retinitis pigmentosa, cotransfection and patch–clamp studies of mutant isoforms with the wild-type bestrophin resulted in Cl conductance that was not significantly different from that measured with the mutant alone, whereas the corresponding experiment in AR retinitis pigmentosa resulted in a significantly greater Cl conductance. 6 Thus, abolished Cl conductance in the retinal pigment epithelium may represent a pathogenetic factor, modulating the phenotype. 
In this article, we focus on vitelliform macular dystrophy associated with mutations in BEST1 that may occur in BMD or in ARB. Typical vitelliform macular dystrophy in association with mutations in BEST1 (AD BMD) usually manifests a vision reduction in the first or second decade of life; however, there is a large variability in expressivity of the disease. 13,14 Typically, it progresses through various clinical stages, leading to increasing central retinal dysfunction with age. 13,14 Most patients preserve visual acuity in at least one eye throughout life. 1,13,14  
To examine the extent of retinopathy and to discover new genotype–phenotype correlations in a larger cohort, patients from families in which at least one family member presented with vitelliform macular dystrophy associated with mutations in BEST1 were identified in two referral centers for hereditary retinal disorders in Denmark and Sweden. 
Affected patients from five Swedish families (harboring the dominant mutations p.Val9Ala, p.Tyr85His, p.Val89Ala, and p.Asp104Glu and the compound heterozygous mutations p.Arg141His/p.Tyr29stop in BEST1) 1,2,7 and four Danish families with vitelliform macular dystrophy, but without a known molecular genetic diagnosis beforehand, were further examined. 
Methods
Subjects
Fifteen Swedish patients (from five different families) out of at least 27 patients with BMD and mutation in BEST1, presenting at the Department of Ophthalmology in Lund, Sweden, since 1988, were examined. 1,2,7 In addition, nine Danish patients from four families presenting since 2007 at the National Eye Clinic for the Visually Impaired, Kennedy Center, Denmark, with a clinical phenotype compatible with BMD, but with no genetic diagnosis, were examined (Fig. 1, Tables 1, 2). Informed consent was obtained from all patients participating in the study or from their legal guardians in accordance with the Declaration of Helsinki. Approval of the study was obtained from an ethics committee. 
Table 1.
 
Clinical Characteristics and Mutations in BEST1 in Swedish Families
Table 1.
 
Clinical Characteristics and Mutations in BEST1 in Swedish Families
Family Patient Sex Age Age at Onset (Age First Documented VA, VA OD/OS) Clinical Findings OD/OS Unifocal/multifocal OD/OS
SI I:1 M 67 Asymptomatic (67, 1.0/1.0) Previtelliform/previtelliform Normal
SI I:2 F 59 Asymptomatic (59, 1.0/1.0) Previtelliform/previtelliform Normal
SI II:1 F 33 10 (10, 0.1/0.4) Atrophic+ fibrotic/atrophic+ fibrotic Unifocal
SI II:2 F 30 4 (4, 0.9/0.1) Vitelliruptive/vitelliruptive Multifocal
SII II:2 M 83 50 (50, 0.1/0.2) Atrophic/atrophic Multifocal
SII III:1 F 46 40 (40, 1.0/0.7) Vitelliform/vitelliruptive Unifocal
SII IV:1 F 19 Asymptomatic (19, 1.0/1.0) Previtelliform/previtelliform Normal
SIII III:1 F 44 13 (13, 1.0/0.7) Vitelliform/fibrotic Unifocal
SIV VII:8 M 51 10 (10, 0.6/0.6) Vitelliruptive/vitelliruptive Multifocal
SIV VI:9 F 75 10 (50, 0.1/0.1) Atrophic+fibrotic/atrophic+fibrotic Unifocal
SIV VII:1 F 49 10 (30, 0.1/0.1) Atrophic/atrophic Multifocal
SIV VII:14 F 32 10 (10, 0.2/0.2) Vitelliruptive/vitelliruptive+ fibrotic Unifocal
SIV VIII:4 M 7 7 (7, 0.2/0.5) Psedohypopyon/fibrotic Unifocal
SV III:1 F 50 30 (30, 0.8/0.8) Vitelliruptive + atrophic/vitelliruptive + atrophic Multifocal
SV IV:1 F 27 10 (10, 0.8/0.8) Vitelliruptive/fibrotic Unifocal/multifocal
Family Patient BEST1 Mutation* Exon Effect OCT Foveal Thickness (OD/OS μm)† VA at Present OD/OS
SI I:1 c.87C>G Exon 2 p.Tyr29stop 165/160 1.0/1.0
SI I:2 c.422G>A Exon 4 p.Arg141His 199/190 1.0/1.0
SI II:1 c.87C>G and c.422G>A Exons 2 and 4 p.Tyr29stop. p.Arg141His 290/280 0.1/0.3
SI II:2 c.87C>G and c.422G>A Exons 2 and 4 p.Tyr29stop. p.Arg141His 406/341 0.1/0.3
SII II:2 c.266T>C Exon 4 p.Val89Ala 110/83 0.1/0.2
SII III:1 c.266T>C Exon 4 p.Val89Ala 232/199 1.0/0.7
SII IV:1 c.266T>C Exon 4 p.Val89Ala 159/199 1.0/1.0
SIII III:1 c.312C>A Exon 4 p.Asp104Glu 298/240 1.0/0.2
SIV VII:8 c.253T>C Exon 4 p.Tyr85His 348/274 0.3/0.1
SIV VI:9 c.253T>C Exon 4 p.Tyr85His 175/133 CF/CF‡
SIV VII:1 c.253T>C Exon 4 p.Tyr85His 398/257 0.1/0.1
SIV VII:14 c.253T>C Exon 4 p.Tyr85His 182/240 0.5/0.9
SIV VIII:4 c.253T>C Exon 4 p.Tyr85His 581/307 0.2/0.5
SV III:1 c.26T>C Exon 2 p.Val9Ala 409/265 0.7/0.5
SV IV:1 c.26T>C Exon 2 p.Val9Ala 282/323 0.9/0.1
Normal (range) 182 (157–207)
Family Patient mfERG§
Rings 1–2 Rings 3–6
Amplitude (nV/deg2) Latency (ms) Amplitude (nV/deg2) Latency (ms)
SI I:1 35.4 27.5 17.4 29.2
SI I:2 27.8 30 9.3 30
SI II:1 5.1 31.6 ND ND
SI II:2 16.2 32.5 17.2 31.7
SII II:2 12.3 30 14.5 30
SII III:1 63.3 29.2 28.7 27.5
SII IV:1 74.6 28.3 32.5 26.7
SIII III:1 26.4 30 28.8 26.7
SIV VII:8 17.1 32.5 22.7 28.3
SIV VI:9 12 33.3 13.9 30
SIV VII:1 15 34.2 18.7 30
SIV VII:14 32.5 27.5 33.3 27.5
SIV VIII:4 Not performed
SV III:1 Not performed
SV IV:1 Not performed
Normal (range)‖ 29.3 (22.8–35.2) 26.3 (25.8–29.5) 12.7 (10.6–20.2) 26.3 (25.0–28.3)
Family Patient EOG Arden Ratio ffERG¶
Rod Rod-Cone 30-Hz-Flicker
Amplitude Amplitude Amplitude Implicit Time
SI I:1 1.5/1.5 104% 86% 92% 110%
SI I:2 2.2/2.0 41% 55% 25% 116%
SI II:1 1.0/1.0 51% 53% 40% 125%
SI II:2 1.0/1.1 104% 52% 106% 115%
SII II:2 1.4/1.4 61% 68% 51% 115%
SII III:1 1.1/1.0 106% 97% 62% 101%
SII IV:1 1.0/1.0 110% 95% 43% 96%
SIII III:1 1.4/1.2 100% 80% 96% 101%
SIV VII:8 1.2/1.2 111% 69% 130% 109%
SIV VI:9 1.0/1.1 65% 104% 75% 117%
SIV VII:1 1.0/1.0 156% 100% 70% 113%
SIV VII:14 1.1/1.2 106% 100% 221% 101%
SIV VIII:4 Not performed Not performed
SV III:1 1.0/0.9 142% 130% 66% 107%
SV IV:1 1.3/1.2 142% 105% 58% 105%
Table 2.
 
Clinical Characteristics and Mutations in BEST1 in Danish Families
Table 2.
 
Clinical Characteristics and Mutations in BEST1 in Danish Families
Family Patient Sex Age Age at Onset (Age at First Documented VA, VA OD/OS) Clinical Findings OD/OS Unifocal/Multifocal
DI III:1 M 41 41 (41, 0.8/0.8) Vitelliruptive/vitelliruptive Multifocal
DI III:2 F 38 Childhood (38 0.1/1.0) Vitelliruptive + atrophic + fibrotic/vitelliruptive Multifocal
DI IV:1 M 13 7 (7, 0.05/0.7) Atrophic + fibrotic/vitelliruptive Unifocal
DII II:1 M 49 20 (39, 0.3/0.5) Atrophic/atrophic Unifocal
DII III:7 M 9 9 (9, 0.5/0.4) Vitelliform/vitelliform Multifocal
DIII I:1 M 40 Asymptomatic (40, 1.0/1.0) Previtelliform/previtelliform Normal
DIII II:1 M 6 6 (6, 0.8/0.8) Vitelliform/vitelliform Unifocal
DIV II:1 F 57 40 (47, 0.6/0.9) Vitelliruptive + atrophic/vitelliruptive Multifocal
DIV II:2 F 44 30 (30, 0.1/0.6) Vitelliruptive + pseudohypopyon/vitelliruptive + pseudohypopyon Unifocal
Family Patient BEST1 Mutation Exon Effect OCT Foveal Thickness (μm) Spectral Domain OCT Foveal IS/OS Junction OD/OS VA at Present OD/OS
DI III:1 c.253T>C Exon 4 p.Tyr85His 176/185 Normal/focally disrupted 0.8/0.8
DI III:2 c.253T>C Exon 4 p.Tyr85His 115/108 Absent/focally disrupted 0.1/1.0
DI IV:1 c.253T>C Exon 4 p.Tyr85His 110/156 Absent/normal 0.05/1.0
DII II:1 c.936C>A Exon 8 p.Asp312Glu 61/76 Absent/absent 0.2/0.6
DII III:7 c.936C>A* Exon 8 p.Asp312Glu* 155/244 Normal/focally disrupted 0.5/0.4
DIII I:1 c.275G>A Exon 4 p.Arg92His 158/173 Normal/normal 1.0/1.0
DIII II:1 c.275G>A Exon 4 p.Arg92His 95/155 Focally disrupted/focally disrupted 0.8/0.8
DIV II:1 c.244C>G Exon 3 p.Leu82Val 208/174 Focally disrupted/focally disrupted 0.9/0.5
DIV II:2 c.244C>G Exon 3 p.Leu82Val 323/315 Absent/focally disrupted 0.2/0.3
Normal (range) 182 (157–207)
Family Patient mfERG
Rings 1–2 Rings 3–6
Amplitude (nV/deg2) Latency (ms) Amplitude (nV/deg2) Latency (ms)
DI III:1 30.2 31.7 16.1 31.7
DI III:2 21 30.8 20.5 29.2
DI IV:1 15.4 27.5 26.7 29.2
DII II:1 5.5 30.8 12.9 30
DII III:7 Not performed
DIII I:1 46.4 29.2 20.5 28.3
DIII II:1 Not performed
DIV II:1 20.8 28.3 16.1 29.2
DIV II:2 12.9 26.7 13.5 27.5
Normal (range) 29.3 (22.8–35.2) 26.3 (25.8–29.5) 12.7 (10.6–20.2) 26.3 (25.0–28.3)
Family Patient EOG Arden Ratio ffERG†
Rod Rod-Cone 30-Hz Flicker
Amplitude Amplitude Amplitude Implicit Time
DI III:1 1.1/1.0 104% 90% 63% 103%
DI III:2 1.2/1.1 Not performed
DI IV:1 1.1/1.1 143% 110% 129% 94%
DII II:1 1.4/1.5 108% 74% 64% 110%
DII III:7 Not performed 35% 50% 60% 91%
DIII I:1 1.4/1.4 153% 91% 90% 97%
DIII II:1 Not performed Not performed
DIV II:1 1.57/0.93 108% 120% 75% 104%
DIV II:2 1.87/1.85 84% 98% 66% 112%
Clinical Examination
Full-field ERG was recorded in an analysis system (Nicolet Biomedical Instruments, Madison, WI), as described previously, in Swedish families and in Danish family DIV. 15 Briefly, subjects' right eyes were dark adapted for 30 minutes before electroretinography. A commercial system (Nicolet Biomedical Instruments, Madison, WI) was used to record ffERG after the pupil was dilated with topical 1% cyclopentolate and 2.5% phenylephrine. After topical anesthesia of the eye, a Burian-Allen bipolar contact lens was applied on the cornea, and a ground electrode was applied to the forehead. Responses were obtained with a wide-band filter (−3 dB at 1 Hz and 500 Hz) stimulated with a single full-field flash (30 μs) if blue light (Wratten filter 47, 47A, and 47B; Eastman Kodak, Rochester, NY) and white light (3.93 [cd · s]/m2). Cone responses were obtained with 30-Hz flickering white light (0.81 [cd · s]/m2) averaged from 20 sweeps. In the Danish families, except family DIV, ffERGs were performed according to the standardized ISCEV (International Society for Clinical Electrophysiology of Vision) protocol. 16 To enable comparison responses between patients, the data are presented as a percentage of the median of normal (56 and 85 normal subjects, respectively, for the two procedures). 
Multifocal electroretinograms were recorded in patients' right eyes (except in SIII III:1 and DI III:1, in whom recordings were obtained from the left eyes) with a visual evoked response imaging system (VERIS 4; EDI, San Mateo, CA), as described previously. 15 Responses from rings 1 and 2 were averaged as 1–2 and responses from rings 3 to 6 were averaged as 3–6. 
OCT (OCT-3 in Swedish families and OCT-4 in Danish families; Carl Zeiss Meditec, Dublin, CA) was performed with single-line horizontal scans centered over the fovea. Foveal thickness was measured in 48 eyes of 24 patients, by calipers placed on the highly reflective layers corresponding to the RPE and the junction between the internal limiting membrane and the vitreous. In addition, using OCT-4, a qualitative evaluation of the highly reflective layer corresponding to the junction between the foveal photoreceptor outer and inner segments (IS/OS), was performed in Danish families (18 eyes of nine patients), and three main categories were recognized: (1) a continuous normal IS/OS layer, (2) an IS/OS layer with focal disruptions, and (3) absence of the foveal IS/OS layer. 
EOG was recorded with an analysis system (Nicolet Biomedical Instruments, Madison, WI), as described previously. 1,2,7  
In addition, in Danish patients, fundus autofluorescence (FAF) photography was recorded with a confocal scanning laser ophthalmoscope (Spectralis HRA; Heidelberg Engineering, Heidelberg, Germany). In family DII, fundus morphology was examined with a combined OCT and scanning laser fundus camera (Spectralis HRA-OCT; Heidelberg Engineering). 
The Spearman rank order correlation test was used to calculate significant correlations between (1) mfERG, visual acuity (VA), and retinal thickness and (2) ffERG responses and the EOG Arden ratio. The analysis of variance test was used to assess the relation between the integrity of the IS/OS junction and VA. 
Molecular Genetic Analysis
DNA was extracted from the peripheral blood samples collected from nine family members (families DI through DIV, Table 2). All BEST1 coding exons were amplified by standard PCR followed by direct sequencing of the corresponding fragments. 
In addition, DNA samples of two patients from family DIV were analyzed by a -microarray technique (BEST-BMD APEX; performed at Asper Ophthalmics, Tartu, Estonia), screening for known mutations in BEST1. Finally, after the finding of an unusual phenotype featuring central bull's eye–like lesions in both eyes, a DNA sample from II:1 in family DII was analyzed by a gene chip microarray technique (ABCR-chip, APEX; Asper Ophthalmics), screening for known mutations in ABCA4
Results
BEST1 Mutations in Family Members
Mutation analysis of the index cases in the four Danish families identified four BEST1 missense mutations: c.244C>G (exon 3), c.253T>C (exon 4), c.275G>A (exon 4), and c.936C>A (exon 8) (Table 2). The c.936C>A transversion (Fig. 2A), leading to p.Asp312Glu, is a novel dominant mutation found in family DII, for which the affected father was heterozygous and his affected son was homozygous (Figs. 1, 2A). Amino acid 312 is located in a highly conserved region of bestrophin (Fig. 2B) and is perfectly preserved in different species. Mutations in many amino acids flanking it cause either Best disease, adult-onset VMD, or ARB. Furthermore, a pathologically reduced EOG Arden ratio in the presence of a normal ffERG in the heterozygous father in family DII indicates that p.Asp312Glu is a disease-causing mutation leading to AD Best disease. 
Figure 1.
 
Pedigrees of Danish and Swedish families with BEST1 mutations. Complete pedigrees for Swedish families have been described by us elsewhere 1,2,7 and thus are outlined here only briefly. Recruited affected individuals are marked with numbers in the different pedigrees. M/+ denotes heterozygosity for mutations in BEST1, whereas M/M denotes homozygosity and M1/M2 denotes compound heterozygosity. No clinical or genetic information was available regarding the relatives of II:1 and III:1 of family DII, the information regarding their family members was obtained by history.
Figure 1.
 
Pedigrees of Danish and Swedish families with BEST1 mutations. Complete pedigrees for Swedish families have been described by us elsewhere 1,2,7 and thus are outlined here only briefly. Recruited affected individuals are marked with numbers in the different pedigrees. M/+ denotes heterozygosity for mutations in BEST1, whereas M/M denotes homozygosity and M1/M2 denotes compound heterozygosity. No clinical or genetic information was available regarding the relatives of II:1 and III:1 of family DII, the information regarding their family members was obtained by history.
Figure 2.
 
A novel AD mutation in the BEST1 gene. (A) The chromatograms of the BEST1 region including the mutated nucleotide (c.936C>A) are presented with a normal sequence (top), the affected heterozygous father (II:1; middle), and the affected homozygous son (III:1, bottom). (B) An amino acid sequence alignment of bestrophin1 (amino acids 287-337 in the human protein) from different species. Amino acids in which mutations were reported to cause a retinal disease are highlighted (light blue, Best disease; green, adult onset VMD and ARB; purple, ARB). Amino acid 312 is highly preserved and marked within a box.
Figure 2.
 
A novel AD mutation in the BEST1 gene. (A) The chromatograms of the BEST1 region including the mutated nucleotide (c.936C>A) are presented with a normal sequence (top), the affected heterozygous father (II:1; middle), and the affected homozygous son (III:1, bottom). (B) An amino acid sequence alignment of bestrophin1 (amino acids 287-337 in the human protein) from different species. Amino acids in which mutations were reported to cause a retinal disease are highlighted (light blue, Best disease; green, adult onset VMD and ARB; purple, ARB). Amino acid 312 is highly preserved and marked within a box.
No mutations were found in ABCA4 in patient II:1 from family DII. 
Ophthalmic Examination
Most patients showed typical features of BMD, as was appreciated by funduscopy in the Swedish families (Fig. 3) and by funduscopy and FAF of patients from the Danish families (Fig. 4). FAF findings included hyperautofluorescence corresponding to vitelliform alterations in the central and paracentral retina. A multifocal pattern was evident in a few patients, including a 9-year-old boy homozygous for the BEST1 p.Asp312Glu mutation (described later) and in whom the changes found on fundus autofluorescence photography exceeded those shown by ophthalmoscopy. Atrophy corresponded to hypoautofluorescence, most evident in a 49 year-old patient years from consanguineous family DII, who harbored the novel mutation p.Asp312Glu heterozygously. 
Figure 3.
 
Fundus appearance, OCT, and mfERGs in representative cases from each Swedish family. Typical central reduction of mfERG responses in all affected patients is circled in the mfERG plot from family SI II:2.
Figure 3.
 
Fundus appearance, OCT, and mfERGs in representative cases from each Swedish family. Typical central reduction of mfERG responses in all affected patients is circled in the mfERG plot from family SI II:2.
Figure 4.
 
Fundus appearance, OCT, and mfERGs in patients from Danish families affected with vitelliform macular dystrophy associated with mutations in BEST1. Multifocal hyperautofluorescence extends beyond the visible vitelliform alterations in, for example, subjects DI III:1 and DII III:7.
Figure 4.
 
Fundus appearance, OCT, and mfERGs in patients from Danish families affected with vitelliform macular dystrophy associated with mutations in BEST1. Multifocal hyperautofluorescence extends beyond the visible vitelliform alterations in, for example, subjects DI III:1 and DII III:7.
Optical coherence tomography (OCT) presented variable patterns (Tables 1, 2, Figs. 3, 4), according to clinical stage of disease, ranging from normal findings (e.g., patient DIII I:1 in Fig. 4) to cystoid macular edema and thickening of the outer retina–choroid complex (ORCC; e.g., patient SI II:2 in Fig. 3 and patient DIII III:1 in Fig. 4, respectively). Retinal thinning and atrophy was seen in the older patients and in one patient aged 49 years from consanguineous family DII, who harbored the novel mutation p.Asp312Glu heterozygously. 
A reduced EOG Arden ratio is considered to be the most penetrant clinical sign of BMD. All patients had pathologic EOG Arden ratios except for SI I:2 (heterozygous for p.Arg141His) and DIV I:2 (heterozygous for p.Leu82Val). 
Of note, the ffERGs were abnormal in five individuals: two unrelated patients (SII II:2 and SIV VI:9 Table 1) and three individuals from family SI, reported by us previously. 7  
mfERG typically revealed central reduction of function and relatively preserved function in the periphery (Tables 1, 2, Figs. 3, 4). However, even in the presence of structural pathology on OCT (thickening of the ORCC marked by a red arrow in Fig. 5), vision and mfERG amplitudes may be preserved, as in individual SII III:1 who is heterozygous for the p.Val89Ala mutation (Fig. 5). 
Figure 5.
 
Patient III:1 in family SII (heterozygous for the Val89Ala mutation) has supranormal mfERG response and localized thickening of the ORCC and subretinal fluid on OCT (arrow).
Figure 5.
 
Patient III:1 in family SII (heterozygous for the Val89Ala mutation) has supranormal mfERG response and localized thickening of the ORCC and subretinal fluid on OCT (arrow).
Family DII exhibits an extremely rare case in which a patient is homozygous for a dominant mutation due to consanguinity. Moreover, we were able to clinically evaluate both the heterozygous affected father and his homozygous affected son. Sequencing analysis of the BEST1 gene in the index patient revealed a heterozygous novel mutation, c.936C>A, leading to p.Asp312Glu. To better characterize the retinal structure of these patients, we performed HRA-OCT. In patient II:1, there was an abrupt termination of the photoreceptor layer in the transition between the area of normal FAF and the area of reduced FAF (Fig. 6, green arrow on OCT). In his homozygous son, the outer retinal layers were better preserved, albeit of reduced reflectivity because of the bowing effect from vitelliform subretinal accumulated material (Fig. 6). The ffERG rod response was reduced in the homozygous boy, indicating widespread retinal degeneration, whereas the heterozygous father had a normal ffERG. No other relatives were available for clinical or genetic evaluation (Fig. 1). 
Figure 6.
 
HRA-OCT reveals loss of the photoreceptor layer (green arrows indicate abrupt transition) in central and peripheral scans in the father, heterozygous for the c.936C>A mutation, leading to p.Asp312Glu, in family DII. The homozygous son presented with a multifocal vitelliform retinopathy. The photoreceptor layer was better preserved over the vitelliform lesion, albeit of reduced reflectivity due to the bowing from the subretinal accumulation of vitelliform material. Yellow areas indicate the extent of the horizontal OCT strip.
Figure 6.
 
HRA-OCT reveals loss of the photoreceptor layer (green arrows indicate abrupt transition) in central and peripheral scans in the father, heterozygous for the c.936C>A mutation, leading to p.Asp312Glu, in family DII. The homozygous son presented with a multifocal vitelliform retinopathy. The photoreceptor layer was better preserved over the vitelliform lesion, albeit of reduced reflectivity due to the bowing from the subretinal accumulation of vitelliform material. Yellow areas indicate the extent of the horizontal OCT strip.
To better characterize the visual function, we performed correlation analysis of mfERG amplitude and implicit time versus VA (Tables 1, 2). mfERG P1 amplitudes and implicit times from rings 1 and 2 correlated significantly with VA (P = 0.017, r = 0.54 and P = 0.012, r = 0.56, respectively, Figs. 7, 8). There was no significant correlation between foveal thickness, as measured with OCT, and VA or mfERG P1 amplitudes from rings 1 and 2 (P = 0.243 [logMAR VA] and P = 0.822 [mfERG], respectively). mfERG latencies were prolonged, for example, in patient SII II:2, in whom end-stage atrophy and thinning of the retina were evident on OCT (Table 2, Fig. 3), and in patient DI I:1 who had an extended distribution of hyperautofluorescence on fundus autofluorescence photography. No significant correlation between VA and age was detected (P = 0.221, logMAR VA) and a few young patients had severely reduced VA (Fig. 9). 
Figure 7.
 
Scatterplot showing a significant correlation between mfERG P1 amplitudes from rings 1 and 2 and VA.
Figure 7.
 
Scatterplot showing a significant correlation between mfERG P1 amplitudes from rings 1 and 2 and VA.
Figure 8.
 
Scatterplot showing a significant correlation between mfERG P1 latencies from rings 1 and 2 and VA.
Figure 8.
 
Scatterplot showing a significant correlation between mfERG P1 latencies from rings 1 and 2 and VA.
Figure 9.
 
The relationship between VA in the better eye and age. Driving vision was maintained in most patients until at least 60 years of age; however, a few young patients had significant visual disability.
Figure 9.
 
The relationship between VA in the better eye and age. Driving vision was maintained in most patients until at least 60 years of age; however, a few young patients had significant visual disability.
Analysis of the mfERG amplitude (Fig. 10) and implicit time (Fig. 11) showed that the amplitude reduction was limited to the central retina, whereas the implicit time was delayed in both the central and peripheral retina in several patients. OCT-4 was performed in both eyes in patients from Danish families, enabling analysis of retinal sublayers in 18 eyes of 9 patients. There was a significant correlation between the degree of preservation of the integrity of the IS/OS layer and VA (P = 0.026, r = 0.72; Fig. 12). 
Figure 10.
 
mfERG amplitudes from center (rings 1–2) and periphery (rings 3–6) showing central reduction of function, but peripheral amplitudes were preserved in most patients.
Figure 10.
 
mfERG amplitudes from center (rings 1–2) and periphery (rings 3–6) showing central reduction of function, but peripheral amplitudes were preserved in most patients.
Figure 11.
 
mfERG latencies from the center (rings 1–2) and periphery (rings 3–6) showing delays not only in the center, but also in the periphery.
Figure 11.
 
mfERG latencies from the center (rings 1–2) and periphery (rings 3–6) showing delays not only in the center, but also in the periphery.
Figure 12.
 
The significant correlation between the integrity of the foveal photoreceptor IS/OS junction by OCT-4 and VA.
Figure 12.
 
The significant correlation between the integrity of the foveal photoreceptor IS/OS junction by OCT-4 and VA.
There was no significant correlation between ffERG response amplitude or implicit time and the EOG Arden ratio (P > 0.13 for all responses). 
Discussion
The diminished EOG light peak is believed to be caused by a defective Ca2+-activated Cl channel in the basolateral membrane of the retinal pigment epithelium (RPE), where bestrophin is expressed. 10 The disruption of ion flow across the membrane due to BEST1 mutations may explain the pathologic changes demonstrated in the pathologic EOG Arden ratio in all but two patients in the present study. If so, since bestrophin is expressed throughout the RPE, one might expect a correlation between the magnitude of the Arden ratio and the total retinal function; however, this was not the case. Furthermore, most patients had a normal ffERG. A disproportionate reduction of the EOG compared to the ffERG indicates a primary defect in the function of the RPE. The generalized retinal degeneration shown by a reduced ffERG in patients with biallelic BEST1 mutations—for example, in families SI and DII—may result from a mechanism different from the one that causes macular degeneration. 
A reduced EOG Arden ratio is considered to be the most penetrant clinical sign of BMD. In the present study, all patients except for SI I:2 (heterozygous for p.Arg141His) and DIV I:2 (heterozygous for p.Leu82Val) had clearly reduced EOG Arden ratios. Patient DIV I:2 demonstrated a borderline reduction of the Arden ratio, and clinical signs including mfERG, OCT, and FAF compatible with BMD. Moreover, the patients' sister harbored the same mutation, and had a reduced EOG Arden ratio (Table 2). The mutation was also described previously in a patient with BMD in a study that included EOG; however, EOG details were not given for this particular patient. 17  
Regarding p.Arg141His, a normal EOG was similarly described in one clinically unaffected individual heterozygous for this mutation, whose daughter carried the compound heterozygous mutations p.Leu41Pro/p.Arg141His in BEST1 and was affected with ARB. 4 Thus, the mutation may not be penetrant in the EOG Arden ratio and seems to cause disease only in a compound heterozygous or recessive mode. 
In this study, we found a novel mutation in exon 8, c.936C>A leading to p.Asp312Glu. The mutation is located in a conserved protein region (Fig. 2B) and several BMD-causing mutations have been reported to affect adjacent amino acids at positions 292 to 311 (see Sohn et al. 3 for a report regarding the p.Glu292Lys mutation and a review by White et al. 18 ). A mutation causing adult-onset VMD and affecting the same amino acid, p.Asp312Asn, has been reported previously. 19 The latter was also reported as a compound heterozygous mutation in association with autosomal recessive bestrophinopathy, together with the mutation p.Met325Thr. 4 In addition, in a recent report, mutations affecting the N terminus (p.Arg19Cys, p.Arg25Cys, and p.Lys30Cys) and the C terminus (p.Gly299Glu, p.Asp301Asn, and p.Asp312Asn) caused channel dysfunction, probably resulting from disruption of the N and C terminus interaction. 20 We therefore conclude that p.Asp312Glu is a probable disease-causing mutation. Consanguineous marriages in this family led to a rare situation in which a patient was homozygous for a disease-causing mutation. Of interest, the patient presented with a multifocal VMD and evidence of widespread retinal degeneration as demonstrated by ffERG. The heterozygous father of the homozygous boy, however, demonstrated atypical Bull's eye–like central atrophy in both eyes. The heterozygous father presented with ffERG responses within normal limits and had a clearly reduced EOG Arden ratio, compatible with AD BMD, which provides further support for the notion that this novel mutation is pathogenic. 
Histopathology from a patient homozygous for the dominant mutation p.Trp93Cys and another patient heterozygous for the mutation p.Thr6Arg demonstrated a similar increase in the retinal pigment epithelium content of A2E and lipofuscin in both patients. The homozygous patient did not demonstrate any more severe structural alterations than the heterozygous patient. 21 This finding seems to differ from those in a homozygous patient in the present study who demonstrated a multifocal vitelliform dystrophy and had signs of widespread retinal degeneration featuring a reduced ffERG rod response. A similar reduction of ffERG rod response was reported previously in ARB. 4  
Different mutations may cause disease by different mechanisms. A functional analysis of an exon 8 variant (p.Gln293His) in human embryonic kidney cells revealed a severe reduction of chloride current that behaved in a dominant negative manner, inhibiting the function of wild-type bestrophin-1 channels. 22 In the present study, the finding of a homozygous dominant mutation in a patient DII II:1 with vitelliform macular dystrophy and evidence of widespread retinal degeneration, may indicate that the pathogenesis of the macular degeneration differs from that of the generalized retinal degeneration. 
Delayed implicit time in the ffERG 30-Hz cone flicker, indicating widespread retinal degeneration, was seen in two sisters from family SI aged 30 and 33 years, who harbored compound heterozygous mutations in BEST1. 7 The genetic alteration in the former seems to closely resemble ARB and those found in canine multifocal retinopathy, which has been described as a recessive disease, with the mutations p.Arg25X and p.Gly161Asp in best1. 4,8 In ARB, patch–clamp studies showed a reduced channel function that was restored after cotransfection with wild-type bestrophin, consistent with a loss-of (channel)-function mechanism of disease. 4 However, biological events other than regulation of ion flow in the retinal pigment epithelium, such as ceramide accumulation, may be involved in the bestrophin-associated disease process. 12  
Based on the findings in the present study and previous studies, it seems that compound heterozygous, biallelic recessive or homozygous dominant mutations in BEST1 may confer a particularly severe phenotype, featuring widespread retinal degeneration, in addition to VMD. 4,7 On the other hand, in dominant heterozygous BMD, the variable phenotype is again highlighted in the present study, and there seems to be no clear pattern relating type of BEST1 mutation to severity of clinical expression. For example, in family DIII with the heterozygous BEST1 mutation c.275G>A leading p.Arg92His, a 6-year-old boy presented with early-onset VMD, including reduced vision and thickening of the outer retina, whereas the only significant finding in the father of the boy was a reduced EOG Arden ratio (Fig 4). Similar intrafamilial phenotypic variability was demonstrated, for example, in family DI, with the heterozygous mutation c.253T>C leading to p.Tyr85His and in family SII with the heterozygous mutation c.266T>C leading to p.Val89Ala. On the other hand, a bilateral multifocal VMD was seen in at least four of eight individuals with the former genotype (c.253T>C leading to p.Tyr85His; Table 1, 2; Figs. 3, 4). Thus, multifocal VMD may be overrepresented in this genotype. The severity of the novel mutation in exon 8, c.936C>A leading to p.Asp312Glu, found in the large consanguineous family DII remains to be determined, as only one heterozygous individual was available for examination. 
mfERG typically revealed amplitude reduction in the central macular area and preserved function in the periphery (Figs. 3, 4). This finding has been reported previously in BMD, and may be a feature in common with other disorders affecting primarily the retinal pigment epithelium, such as Bothnia dystrophy. 23 However, a few patients had preserved mfERG responses, normal visual acuities, and normal OCT scans, consistent with variable expression and penetrance of disease phenotype. mfERG amplitudes were significantly correlated to VA, which is in line with previous results. 24 mfERG implicit times, however, were delayed in the periphery in several patients, indicating a subtle dysfunction, as well in the periphery, in addition to the central retinal dysfunction. A significant correlation between age and VA has been described in BMD, with most patients younger than 40 years having VA > 20/40. 13 In this study, however, a few young patients had significant visual disability (Fig. 9). 
mfERG implicit times may be a sensitive tool for detecting early functional disturbances in BMD, as has been suggested in diabetic retinopathy and Stargardt disease, in which implicit time delays may predict future sites of retinopathy. 2527 In patients with multifocal distribution of hyperautofluorescence by FAF, peripheral mfERG implicit times were delayed (Figs. 4, 11). In a previous study on mfERG responses in BMD, only a slight but significant delay was noted for peripheral, but not central, implicit times. 24 In the latter, a different form of mfERG was used featuring only 61 hexagonal stimulus elements, compared with 103 in our study, with a decrease in resolution of the responses. 
Different patterns were identified with OCT, according to clinical stage. Some of these, including thickening of the ORCC and subretinal fluid, have been described earlier in BMD. 7,28 The thickened hyperreflective ORCC would probably correspond to the accumulation of lipofuscin and A2E in and beneath the RPE, as has been shown in histopathologic studies of BMD. 21,29 This finding is also supported by the hyperautofluorescence in fundus autofluorescence photography. The accumulation of lipofuscin and A2E is probably one of the primary steps in the pathogenesis of retinal degeneration and visual loss. Despite this pattern on OCT, (e.g., III:1 in family SII p.Val89Ala, Table 2, Fig. 5) VA and central retinal function can still be preserved on mfERG. Recently, however, it has been suggested that a supranormal mfERG is a sign of impending visual loss (Koozekanani DD, et al. IOVS 2006;47:ARVO E-Abstract 3754). 
Fundus autofluorescence photography is a sensitive method of revealing lipofuscin-containing material in various degenerative retinal conditions, including BMD. 3,17,30 The distribution of hyperautofluorescence may extend the funduscopically visible changes, as in patient DI I:1, heterozygous for the mutation p.Tyr85His and in the 9-year-old boy homozygous for the mutation c.936C>A leading to p.Asp312Glu. We have recently shown that patients with fundus albipunctatus due to mutations in RDH5 and hence an inhibition of the retinoid cycle leading to diminished A2E formation have a generalized homogenous reduction of fundus autofluorescence, implying reduced lipofuscin formation. These patients may have relatively well-preserved retinal function, 31 which implies a potential treatment for vitelliform dystrophy and BMD: a pharmacologic inhibition of the retinoid cycle. 32 If so, high-resolution OCT findings—for example, demonstration of preserved photoreceptors and absence of atrophy and fibrosis—may indicate eligibility for treatment. 
Central foveal thickness, as measured with OCT, did not significantly correlate with VA in our study. This finding is similar to retinitis pigmentosa, but in contrast to diabetic retinopathy, where retinal function seems to deteriorate when foveal retinal thickness exceeds 280 μm. 33,34 In Stargardt macular degeneration, a statistically significant association between central foveal thickness and VA has been demonstrated, reflecting the pathogenesis of visual loss in this form of macular degeneration, which seems to be related to progressive central retinal atrophy. 35 In BMD, the pathogenesis is more complex and dynamic, where different types of structural alterations, including thickening of the ORCC and subretinal fluid/retinal edema, precede the end-stage atrophy, thus making it difficult to relate simply foveal thickness to function. In this context, we should mention that the OCT protocol differed among Danish and Swedish patients, which may be a source of bias when seeking to compare retinal thickness among patients. To reduce dissimilarities due to differences in methodology, retinal thickness was measured with calipers in all patients, thus avoiding any bias due to different automated software procedures measuring retinal thickness. Even so, we cannot exclude a systematic bias involved. However, since a comparison between the two groups was not of interest, only a possible association between thickness and VA in the total cohort, this seems to further reduce the influence of a potential bias. 
Furthermore, by OCT-4 which was performed in all Danish patients, a preserved integrity of the foveal photoreceptor IS/OS junction was related to VA, implying a secondary degeneration of foveal photoreceptors in VMD. Similar OCT findings were demonstrated in a previous study in VMD where BEST1 mutations were reported in only a minority of patients. 36  
To conclude, in vitelliform macular dystrophy associated with mutations in BEST1, the combination of molecular genetics, electrophysiology, and OCT is useful in assessment of diagnosis and disease severity. These methods may also provide a suitable tool for evaluating possible treatment outcomes in the future. 
Footnotes
 Supported by grants from the Swedish Society of Medicine, Dag Lenards fond, Stiftelsen för synskadade i f d Malmöhus län, Stiftelsen Kronprincessan Margaretas arbetsnämnd för synskadade, the Skane County Research and Development Fund, The Velux Foundation, The John and Birthe Meyer Foundation, the Danish Research Council, and Grant 3000003241 from the Chief Scientist Office of the Ministry of Health, Israel.
Footnotes
 Disclosure: P. Schatz, None; H. Bitner, None; B. Sander, None; S. Holfort, None; S. Andreasson, None; M. Larsen, None; D. Sharon, None
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Figure 1.
 
Pedigrees of Danish and Swedish families with BEST1 mutations. Complete pedigrees for Swedish families have been described by us elsewhere 1,2,7 and thus are outlined here only briefly. Recruited affected individuals are marked with numbers in the different pedigrees. M/+ denotes heterozygosity for mutations in BEST1, whereas M/M denotes homozygosity and M1/M2 denotes compound heterozygosity. No clinical or genetic information was available regarding the relatives of II:1 and III:1 of family DII, the information regarding their family members was obtained by history.
Figure 1.
 
Pedigrees of Danish and Swedish families with BEST1 mutations. Complete pedigrees for Swedish families have been described by us elsewhere 1,2,7 and thus are outlined here only briefly. Recruited affected individuals are marked with numbers in the different pedigrees. M/+ denotes heterozygosity for mutations in BEST1, whereas M/M denotes homozygosity and M1/M2 denotes compound heterozygosity. No clinical or genetic information was available regarding the relatives of II:1 and III:1 of family DII, the information regarding their family members was obtained by history.
Figure 2.
 
A novel AD mutation in the BEST1 gene. (A) The chromatograms of the BEST1 region including the mutated nucleotide (c.936C>A) are presented with a normal sequence (top), the affected heterozygous father (II:1; middle), and the affected homozygous son (III:1, bottom). (B) An amino acid sequence alignment of bestrophin1 (amino acids 287-337 in the human protein) from different species. Amino acids in which mutations were reported to cause a retinal disease are highlighted (light blue, Best disease; green, adult onset VMD and ARB; purple, ARB). Amino acid 312 is highly preserved and marked within a box.
Figure 2.
 
A novel AD mutation in the BEST1 gene. (A) The chromatograms of the BEST1 region including the mutated nucleotide (c.936C>A) are presented with a normal sequence (top), the affected heterozygous father (II:1; middle), and the affected homozygous son (III:1, bottom). (B) An amino acid sequence alignment of bestrophin1 (amino acids 287-337 in the human protein) from different species. Amino acids in which mutations were reported to cause a retinal disease are highlighted (light blue, Best disease; green, adult onset VMD and ARB; purple, ARB). Amino acid 312 is highly preserved and marked within a box.
Figure 3.
 
Fundus appearance, OCT, and mfERGs in representative cases from each Swedish family. Typical central reduction of mfERG responses in all affected patients is circled in the mfERG plot from family SI II:2.
Figure 3.
 
Fundus appearance, OCT, and mfERGs in representative cases from each Swedish family. Typical central reduction of mfERG responses in all affected patients is circled in the mfERG plot from family SI II:2.
Figure 4.
 
Fundus appearance, OCT, and mfERGs in patients from Danish families affected with vitelliform macular dystrophy associated with mutations in BEST1. Multifocal hyperautofluorescence extends beyond the visible vitelliform alterations in, for example, subjects DI III:1 and DII III:7.
Figure 4.
 
Fundus appearance, OCT, and mfERGs in patients from Danish families affected with vitelliform macular dystrophy associated with mutations in BEST1. Multifocal hyperautofluorescence extends beyond the visible vitelliform alterations in, for example, subjects DI III:1 and DII III:7.
Figure 5.
 
Patient III:1 in family SII (heterozygous for the Val89Ala mutation) has supranormal mfERG response and localized thickening of the ORCC and subretinal fluid on OCT (arrow).
Figure 5.
 
Patient III:1 in family SII (heterozygous for the Val89Ala mutation) has supranormal mfERG response and localized thickening of the ORCC and subretinal fluid on OCT (arrow).
Figure 6.
 
HRA-OCT reveals loss of the photoreceptor layer (green arrows indicate abrupt transition) in central and peripheral scans in the father, heterozygous for the c.936C>A mutation, leading to p.Asp312Glu, in family DII. The homozygous son presented with a multifocal vitelliform retinopathy. The photoreceptor layer was better preserved over the vitelliform lesion, albeit of reduced reflectivity due to the bowing from the subretinal accumulation of vitelliform material. Yellow areas indicate the extent of the horizontal OCT strip.
Figure 6.
 
HRA-OCT reveals loss of the photoreceptor layer (green arrows indicate abrupt transition) in central and peripheral scans in the father, heterozygous for the c.936C>A mutation, leading to p.Asp312Glu, in family DII. The homozygous son presented with a multifocal vitelliform retinopathy. The photoreceptor layer was better preserved over the vitelliform lesion, albeit of reduced reflectivity due to the bowing from the subretinal accumulation of vitelliform material. Yellow areas indicate the extent of the horizontal OCT strip.
Figure 7.
 
Scatterplot showing a significant correlation between mfERG P1 amplitudes from rings 1 and 2 and VA.
Figure 7.
 
Scatterplot showing a significant correlation between mfERG P1 amplitudes from rings 1 and 2 and VA.
Figure 8.
 
Scatterplot showing a significant correlation between mfERG P1 latencies from rings 1 and 2 and VA.
Figure 8.
 
Scatterplot showing a significant correlation between mfERG P1 latencies from rings 1 and 2 and VA.
Figure 9.
 
The relationship between VA in the better eye and age. Driving vision was maintained in most patients until at least 60 years of age; however, a few young patients had significant visual disability.
Figure 9.
 
The relationship between VA in the better eye and age. Driving vision was maintained in most patients until at least 60 years of age; however, a few young patients had significant visual disability.
Figure 10.
 
mfERG amplitudes from center (rings 1–2) and periphery (rings 3–6) showing central reduction of function, but peripheral amplitudes were preserved in most patients.
Figure 10.
 
mfERG amplitudes from center (rings 1–2) and periphery (rings 3–6) showing central reduction of function, but peripheral amplitudes were preserved in most patients.
Figure 11.
 
mfERG latencies from the center (rings 1–2) and periphery (rings 3–6) showing delays not only in the center, but also in the periphery.
Figure 11.
 
mfERG latencies from the center (rings 1–2) and periphery (rings 3–6) showing delays not only in the center, but also in the periphery.
Figure 12.
 
The significant correlation between the integrity of the foveal photoreceptor IS/OS junction by OCT-4 and VA.
Figure 12.
 
The significant correlation between the integrity of the foveal photoreceptor IS/OS junction by OCT-4 and VA.
Table 1.
 
Clinical Characteristics and Mutations in BEST1 in Swedish Families
Table 1.
 
Clinical Characteristics and Mutations in BEST1 in Swedish Families
Family Patient Sex Age Age at Onset (Age First Documented VA, VA OD/OS) Clinical Findings OD/OS Unifocal/multifocal OD/OS
SI I:1 M 67 Asymptomatic (67, 1.0/1.0) Previtelliform/previtelliform Normal
SI I:2 F 59 Asymptomatic (59, 1.0/1.0) Previtelliform/previtelliform Normal
SI II:1 F 33 10 (10, 0.1/0.4) Atrophic+ fibrotic/atrophic+ fibrotic Unifocal
SI II:2 F 30 4 (4, 0.9/0.1) Vitelliruptive/vitelliruptive Multifocal
SII II:2 M 83 50 (50, 0.1/0.2) Atrophic/atrophic Multifocal
SII III:1 F 46 40 (40, 1.0/0.7) Vitelliform/vitelliruptive Unifocal
SII IV:1 F 19 Asymptomatic (19, 1.0/1.0) Previtelliform/previtelliform Normal
SIII III:1 F 44 13 (13, 1.0/0.7) Vitelliform/fibrotic Unifocal
SIV VII:8 M 51 10 (10, 0.6/0.6) Vitelliruptive/vitelliruptive Multifocal
SIV VI:9 F 75 10 (50, 0.1/0.1) Atrophic+fibrotic/atrophic+fibrotic Unifocal
SIV VII:1 F 49 10 (30, 0.1/0.1) Atrophic/atrophic Multifocal
SIV VII:14 F 32 10 (10, 0.2/0.2) Vitelliruptive/vitelliruptive+ fibrotic Unifocal
SIV VIII:4 M 7 7 (7, 0.2/0.5) Psedohypopyon/fibrotic Unifocal
SV III:1 F 50 30 (30, 0.8/0.8) Vitelliruptive + atrophic/vitelliruptive + atrophic Multifocal
SV IV:1 F 27 10 (10, 0.8/0.8) Vitelliruptive/fibrotic Unifocal/multifocal
Family Patient BEST1 Mutation* Exon Effect OCT Foveal Thickness (OD/OS μm)† VA at Present OD/OS
SI I:1 c.87C>G Exon 2 p.Tyr29stop 165/160 1.0/1.0
SI I:2 c.422G>A Exon 4 p.Arg141His 199/190 1.0/1.0
SI II:1 c.87C>G and c.422G>A Exons 2 and 4 p.Tyr29stop. p.Arg141His 290/280 0.1/0.3
SI II:2 c.87C>G and c.422G>A Exons 2 and 4 p.Tyr29stop. p.Arg141His 406/341 0.1/0.3
SII II:2 c.266T>C Exon 4 p.Val89Ala 110/83 0.1/0.2
SII III:1 c.266T>C Exon 4 p.Val89Ala 232/199 1.0/0.7
SII IV:1 c.266T>C Exon 4 p.Val89Ala 159/199 1.0/1.0
SIII III:1 c.312C>A Exon 4 p.Asp104Glu 298/240 1.0/0.2
SIV VII:8 c.253T>C Exon 4 p.Tyr85His 348/274 0.3/0.1
SIV VI:9 c.253T>C Exon 4 p.Tyr85His 175/133 CF/CF‡
SIV VII:1 c.253T>C Exon 4 p.Tyr85His 398/257 0.1/0.1
SIV VII:14 c.253T>C Exon 4 p.Tyr85His 182/240 0.5/0.9
SIV VIII:4 c.253T>C Exon 4 p.Tyr85His 581/307 0.2/0.5
SV III:1 c.26T>C Exon 2 p.Val9Ala 409/265 0.7/0.5
SV IV:1 c.26T>C Exon 2 p.Val9Ala 282/323 0.9/0.1
Normal (range) 182 (157–207)
Family Patient mfERG§
Rings 1–2 Rings 3–6
Amplitude (nV/deg2) Latency (ms) Amplitude (nV/deg2) Latency (ms)
SI I:1 35.4 27.5 17.4 29.2
SI I:2 27.8 30 9.3 30
SI II:1 5.1 31.6 ND ND
SI II:2 16.2 32.5 17.2 31.7
SII II:2 12.3 30 14.5 30
SII III:1 63.3 29.2 28.7 27.5
SII IV:1 74.6 28.3 32.5 26.7
SIII III:1 26.4 30 28.8 26.7
SIV VII:8 17.1 32.5 22.7 28.3
SIV VI:9 12 33.3 13.9 30
SIV VII:1 15 34.2 18.7 30
SIV VII:14 32.5 27.5 33.3 27.5
SIV VIII:4 Not performed
SV III:1 Not performed
SV IV:1 Not performed
Normal (range)‖ 29.3 (22.8–35.2) 26.3 (25.8–29.5) 12.7 (10.6–20.2) 26.3 (25.0–28.3)
Family Patient EOG Arden Ratio ffERG¶
Rod Rod-Cone 30-Hz-Flicker
Amplitude Amplitude Amplitude Implicit Time
SI I:1 1.5/1.5 104% 86% 92% 110%
SI I:2 2.2/2.0 41% 55% 25% 116%
SI II:1 1.0/1.0 51% 53% 40% 125%
SI II:2 1.0/1.1 104% 52% 106% 115%
SII II:2 1.4/1.4 61% 68% 51% 115%
SII III:1 1.1/1.0 106% 97% 62% 101%
SII IV:1 1.0/1.0 110% 95% 43% 96%
SIII III:1 1.4/1.2 100% 80% 96% 101%
SIV VII:8 1.2/1.2 111% 69% 130% 109%
SIV VI:9 1.0/1.1 65% 104% 75% 117%
SIV VII:1 1.0/1.0 156% 100% 70% 113%
SIV VII:14 1.1/1.2 106% 100% 221% 101%
SIV VIII:4 Not performed Not performed
SV III:1 1.0/0.9 142% 130% 66% 107%
SV IV:1 1.3/1.2 142% 105% 58% 105%
Table 2.
 
Clinical Characteristics and Mutations in BEST1 in Danish Families
Table 2.
 
Clinical Characteristics and Mutations in BEST1 in Danish Families
Family Patient Sex Age Age at Onset (Age at First Documented VA, VA OD/OS) Clinical Findings OD/OS Unifocal/Multifocal
DI III:1 M 41 41 (41, 0.8/0.8) Vitelliruptive/vitelliruptive Multifocal
DI III:2 F 38 Childhood (38 0.1/1.0) Vitelliruptive + atrophic + fibrotic/vitelliruptive Multifocal
DI IV:1 M 13 7 (7, 0.05/0.7) Atrophic + fibrotic/vitelliruptive Unifocal
DII II:1 M 49 20 (39, 0.3/0.5) Atrophic/atrophic Unifocal
DII III:7 M 9 9 (9, 0.5/0.4) Vitelliform/vitelliform Multifocal
DIII I:1 M 40 Asymptomatic (40, 1.0/1.0) Previtelliform/previtelliform Normal
DIII II:1 M 6 6 (6, 0.8/0.8) Vitelliform/vitelliform Unifocal
DIV II:1 F 57 40 (47, 0.6/0.9) Vitelliruptive + atrophic/vitelliruptive Multifocal
DIV II:2 F 44 30 (30, 0.1/0.6) Vitelliruptive + pseudohypopyon/vitelliruptive + pseudohypopyon Unifocal
Family Patient BEST1 Mutation Exon Effect OCT Foveal Thickness (μm) Spectral Domain OCT Foveal IS/OS Junction OD/OS VA at Present OD/OS
DI III:1 c.253T>C Exon 4 p.Tyr85His 176/185 Normal/focally disrupted 0.8/0.8
DI III:2 c.253T>C Exon 4 p.Tyr85His 115/108 Absent/focally disrupted 0.1/1.0
DI IV:1 c.253T>C Exon 4 p.Tyr85His 110/156 Absent/normal 0.05/1.0
DII II:1 c.936C>A Exon 8 p.Asp312Glu 61/76 Absent/absent 0.2/0.6
DII III:7 c.936C>A* Exon 8 p.Asp312Glu* 155/244 Normal/focally disrupted 0.5/0.4
DIII I:1 c.275G>A Exon 4 p.Arg92His 158/173 Normal/normal 1.0/1.0
DIII II:1 c.275G>A Exon 4 p.Arg92His 95/155 Focally disrupted/focally disrupted 0.8/0.8
DIV II:1 c.244C>G Exon 3 p.Leu82Val 208/174 Focally disrupted/focally disrupted 0.9/0.5
DIV II:2 c.244C>G Exon 3 p.Leu82Val 323/315 Absent/focally disrupted 0.2/0.3
Normal (range) 182 (157–207)
Family Patient mfERG
Rings 1–2 Rings 3–6
Amplitude (nV/deg2) Latency (ms) Amplitude (nV/deg2) Latency (ms)
DI III:1 30.2 31.7 16.1 31.7
DI III:2 21 30.8 20.5 29.2
DI IV:1 15.4 27.5 26.7 29.2
DII II:1 5.5 30.8 12.9 30
DII III:7 Not performed
DIII I:1 46.4 29.2 20.5 28.3
DIII II:1 Not performed
DIV II:1 20.8 28.3 16.1 29.2
DIV II:2 12.9 26.7 13.5 27.5
Normal (range) 29.3 (22.8–35.2) 26.3 (25.8–29.5) 12.7 (10.6–20.2) 26.3 (25.0–28.3)
Family Patient EOG Arden Ratio ffERG†
Rod Rod-Cone 30-Hz Flicker
Amplitude Amplitude Amplitude Implicit Time
DI III:1 1.1/1.0 104% 90% 63% 103%
DI III:2 1.2/1.1 Not performed
DI IV:1 1.1/1.1 143% 110% 129% 94%
DII II:1 1.4/1.5 108% 74% 64% 110%
DII III:7 Not performed 35% 50% 60% 91%
DIII I:1 1.4/1.4 153% 91% 90% 97%
DIII II:1 Not performed Not performed
DIV II:1 1.57/0.93 108% 120% 75% 104%
DIV II:2 1.87/1.85 84% 98% 66% 112%
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